title: Graded expression of zinc-responsive genes through two regulatory zinc-binding sites in Zur.
abstract: Zinc is one of the essential transition metals in cells. Excess or lack of zinc is detrimental, and cells exploit highly sensitive zinc-binding regulators to achieve homeostasis. In this article, we present a crystal structure of active Zur from Streptomyces coelicolor with three zinc-binding sites (C-, M-, and D-sites). Mutations of the three sites differentially affected sporulation and transcription of target genes, such that C- and M-site mutations inhibited sporulation and derepressed all target genes examined, whereas D-site mutations did not affect sporulation and derepressed only a sensitive gene. Biochemical and spectroscopic analyses of representative metal site mutants revealed that the C-site serves a structural role, whereas the M- and D-sites regulate DNA-binding activity as an on-off switch and a fine-tuner, respectively. Consistent with differential effect of mutations on target genes, zinc chelation by TPEN derepressed some genes (znuA, rpmF2) more sensitively than others (rpmG2, SCO7682) in vivo. Similar pattern of TPEN-sensitivity was observed for Zur-DNA complexes formed on different promoters in vitro. The sensitive promoters bound Zur with lower affinity than the less sensitive ones. EDTA-treated apo-Zur gained its DNA binding activity at different concentrations of added zinc for the two promoter groups, corresponding to free zinc concentrations of 4.5×10(-16) M and 7.9×10(-16) M for the less sensitive and sensitive promoters, respectively. The graded expression of target genes is a clever outcome of subtly modulating Zur-DNA binding affinities in response to zinc availability. It enables bacteria to detect metal depletion with improved sensitivity and optimize gene-expression pattern.
authors.affiliation: Laboratory of Molecular Microbiology, School of Biological Sciences, and Institute of Microbiology, Seoul National University, Seoul 151-742, Republic of Korea.
authors.forename: Jung-Ho
authors.initials: JH
authors.lastname: Shin
authors.forename: Hoi Jong
authors.initials: HJ
authors.lastname: Jung
authors.forename: Young Jun
authors.initials: YJ
authors.lastname: An
authors.forename: Yoo-Bok
authors.initials: YB
authors.lastname: Cho
authors.forename: Sun-Shin
authors.initials: SS
authors.lastname: Cha
authors.forename: Jung-Hye
authors.initials: JH
authors.lastname: Roe
mesh_terms: D001665:Binding Sites; D005029:Ethylenediamines / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000502:physiology*; D005798:Genes, Bacterial / Q000502:physiology*; D009154:Mutation; D020218:Response Elements / Q000502:physiology*; D048372:Streptomyces coelicolor / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*; D015032:Zinc / Q000378:metabolism*
chemical_list: D005029:Ethylenediamines; C099539:tris(2-aminoethyl)amine; D015032:Zinc